We have used restriction analysis, heteroduplex formation, and thermal stability studies to identify and characterize a family of interspersed repetitive sequences from the mouse genome. The repeats are about 400 base pairs long and represent 1-2% of mouse genomic DNA. The sequences are found at variable distances on the 5 feet as well as 3 feet sides of genes. Related sequences are present also in the rat, Syrian hamster, and monkey genomes. Comparison of the sequence of one element with other known repetitive elements has revealed that it is a member of the R-sequence family. Computer analysis of the sequence has revealed an RNA polymerase II promoter, polyadenylation signal, and homology with the enhancer/activator region of the SV40 72 bp repeat. Experiments are in progress to determine whether the R-sequence family members function as promoters of transcription, and whether they can enhance the transcription of sequences under control of other promoters. To look at these questions, we have made constructs containing R-sequence and a vector carrying the bacterial gene chloramphenicol acetyl transferase which upon transfection into mammalian cells can be used in a transient expression assay to measure promoter and enhancer functions.